Duke University?s Superfund Research Center examines the problem of early life exposure to hazardous chemicals and later life consequences. Project 4 addresses this issue in an ecological context, and takes advantage of real world case of multigenerational exposures to mixtures of polycyclic aromatic hydrocarbons (PAHs). An important aspect of long term environmental pollution, often associated with Superfund sites, is that of adaptations by exposed organisms over multiple generations that improve the abilities of populations to thrive in polluted ecosystems but also incur fitness costs. Of particular importance in this regard is the phenomenon of pollution-driven genetic adaptation, that is, the potential for pollution to act as a significant selection pressure potentially driving evolution. This phenomenon of ?evolutionary toxicology? has important ramifications for environmental science and management, including conservation biology, the elucidation of fitness costs, environmental risk assessment, and the evaluation of remediation efforts. Atlantic killifish (Fundulus heteroclitus) inhabiting the Elizabeth River in the Tidewater region of Virginia provide an excellent ?natural experiment? for exploring this phenomenon. Moreover, this system is very relevant to the goals of the Superfund Research Program ? several areas are highly contaminated with mixtures of PAHs emitted by wood treatment plants, including a designated Superfund site, and other contaminants. The Project?s Specific Aims are: 1. To elucidate ?omic differences among Elizabeth River subpopulations of killifish reflecting different PAH exposure histories. 2. To determine associated physiological impacts (e.g. fitness costs) in these subpopulations, including deficits of bioenergetics, aerobic performance and behavior, and in a subset of these populations, evaluate responses to natural stressors (hypoxia and temperature). 3. To determine the persistent and transgenerational effects of PAHs on mitochondrial structure and function and associated organismal impacts using zebrafish and killifish. 4. To monitor temporal responses of selected endpoints in killifish populations from a site undergoing remediation for PAHs, a site high in PAHs not being remediated, and a low PAH reference site. This proposed research project will build upon previous DUSRC research and involve integrated field and laboratory studies utilizing two piscine models ? the Atlantic killifish and zebrafish (Danio rerio). Field studies will center focus on several subpopulations of killifish inhabiting discrete sites in the Elizabeth River that cover a large gradient of PAH contamination, and include sites undergoing remediation. These studies will elucidate relationships among PAH exposures and selected aspects of molecular biology in these populations, as well as their responses to remediation. Laboratory studies will employ offspring of wild- caught killifish to explore fitness costs associated with adaptation to PAH contaminated environments. Studies with both killifish and zebrafish will elucidate impacts of PAH exposures mitochondrial structure and function and associated impacts on aerobic performance and behavior.